Mast cells play important roles in inflammation and in innate immune defense. Upon activation, mast cells release many inflammatory mediators, some of them are preformed and stored in their secretory granules, and are released via regulated exocytosis in a process known as degranulation. The importance of degranulation, different to other mast cell responses, in the development and evolution of inflammation is not known. However, in the past years we have shown that mast cell tryptases released during degranulation play important roles in inflammatory arthritis and defense against bacterial infections. Exocytosis is a highly regulated process and Munc18 proteins are essential components of the exocytic machinery, such that their absence abolishes exocytosis in other cells. Although we know Munc18 proteins are required, the identity of the exocytic step controlled by these proteins remains controversial. Munc18-2 is the main Munc18 isoform in mast cells, and we have created mice with altered activity or expression of Munc18-2 only in their mast cells via genetic manipulation. Preliminary studies of our mutant mice point to a severe and selective defect in mast cell regulated exocytosis. We plan to use these unique reagents to uncover the specifics of Munc18-2 involvement in exocytosis, and to study the participation of mast cell degranulation in immune responses. First, using a combination of biochemical, cellular and electrophysiological assays we propose to determine if mast cell exocytosis is selectively controlled by Munc18-2, and to dissect the specific contribution of this protein that makes it so crucial for exocytosis. Second, using models of allergic asthma, inflammatory arthritis, and of bacterial peritonitis and pneumonia, we plan to determine which important pathophysiological roles of mast cells depend on their degranulation. Our genetically modified mice may reveal if mast cell degranulation is an attractive and safe therapeutic target aimed at controlling the involvement of this cell in inflammatory diseases. PUBLIC HEALTH RELEVANCE: Mast cells play important roles in asthma, rheumatoid arthritis, chronic allergies, and other inflammatory disorders, and in the effective control of bacteria and helminth infections. Using transgenic mice we are manipulating mast cell degranulation selectively and we propose to test the effects of such modification in animal models of allergic asthma, inflammatory arthritis and bacterial infections. If we succeed, we will have identified a potential therapeutic target that either could be enhanced to increase our defenses against infectious organisms or suppressed to treat numerous mast cell-dependent inflammatory disorders.